Many common birth defects are believed to result from defective morphogenesis. Morphogenesis involves cell-cell contacts and cell surface-cytoskeletal interactions. Our long term objective is to elucidate the relationship between cell-cell contact and cell surface cytoskeletal interactions during normal morphogenesis in the mammalian embryo. We propose to study such a relationship as it exists in one of the simplest models available for experiments on mammalian morphogenesis, namely, the formation of the mouse blastocyst in vitro. Blastocyst formation entails two interrelated events, 1) cavitation and 2) differentiation of trophectoderm and inner cell mass (ICM). This project focuses on the morphogenetic details of cavitation. We propose a new model for the onset of cavitation that incorporates the following features: 1. cavitation depends on the cortical localization of cytoplasmic organelles (droplets and mitochondria), which is restricted to apposed cell surfaces of the morula, 2. cortical localization of these organelles to apposed cell surfaces depends on, a. differences in plasma membrane domains between free and apposed cell surfaces, b. interactions between cytoskeletal elements and plasma membrane domains of apposed cell surfaces (i.e., cell surface-cytoskeletal interactions), 3. the droplets consist of lipid and their catabolism by the attendant mitochondria is coupled to the production of nascent blastocoele fluid.
The specific aims of this project are to examine the validity of these proposed features with special attention given to the role of cell surface-cytoskeletal interactions and apposed plasma membrane domains in cortical localization and of lipid catabolism as a source of nascent blastocoele fluid. The experiments will entail the use of antisera (immunofluorescence and immunoperoxidase techniques) that will selectively label specific membrane domains together with treatments that will disrupt cytoskeletal elements, membrane domains and cell surface-cytoskeletal interactions; results will be assessed by light and electron microcopy. Lipid catabolism will be examined by cytochemical, autoradiographic and morphometric analyses at the light and electron microscopic levels.
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